Although it has been established that a loss of synapses occurs in Alzheimer's disease (AD), the neurons responsible for synaptic loss have not been determined. We hypothesize that it is the neurons containing neurofibrillary tangles (NFT) which lose synapses. Neurofibrillary tangles are composed primarily of abnormal aggregation of the protein, tau. It has been speculated that an inability of tau to stabilize microtubules in AD neurons results in a destabilatation of microtubule. The key hypothesis of this proposal is that loss of synapses in AD neurons is ultimately the result of a destabilization of microtubules within the neuron. Since the synaptic endings of neurons in human brain tissue are many centimeters away from their soma, it is not technically possible to trace the length of neurons with axonal transport deficits to their synaptic endings. Therefore, this proposal aims to establish markers that signal neuronal transport loss in AD neurons in postmortem tissue, and to correlate those markers with the loss of synapses within a neuron. Preliminary data form my work has indicated message levels for the synaptic proteins, GAP-43 and synaptphysin, are specifically, and dramatically downregulated in tangle containing neurons relative to neighboring non-tangle neurons. These data suggest the strong likelihood that neurons containing tangles are responsible for the loss of synapses seen in AD, and has led to the hypotheses proposed here. Hypothesis 1: Synapsing primary hippocampal neurons undergoing inhibition of axonal transport exhibit: 1) accumulation of vesicles of vesicles, 2) loss of synapses, and 3) a decrease of synaptic messages, in that order. Specific Aim 1: Establish primary hippocampal cultures and verify the presence of synapses ultrastructurally. Establish the effective dose of nocodozole which results in 1) total and 2) partial loss of microtubules. Verify loss ultrastructurally. Determine the consequence of the loss of microtubules by 1) count synaptophysin, ans synapsis I puncta. Count a subset ultrastructurally, 2) count grains formed in situ hybridization for synaptophysin, synapsis I, Poly A, and cathepsin D. Compare to untreated cultures, 3) count mitochondria and synaptic vesicle in ultrastructural images of axon hillocks. hypothesis 2: Neurons in AD hippocampal tissue contain accumulation of mitochondria at the axon hillock or within the soma, relative to age-matched control neurons. Specific Aim 2: Videocaputure ultrastructural images of AD axon hillock regions. Count mitochondria. Determine percent of NFT occupying neuronal profiles. Compare to controls.